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54 (2) • May 2005: 411–425 Oxelman & al. • Disintegration of

Further disintegration of Scrophulariaceae

Bengt Oxelman1, Per Kornhall1, Richard G. Olmstead2 & Birgitta Bremer3

1 Department of Systematic Botany, Evolutionary Biology Centre, Uppsala University, Norbyvägen 18D SE- 752 36 Uppsala, Sweden. [email protected]. (author for correspondence), [email protected] 2 Department of Biology, Box 355325, University of Washington, Seattle, Washington 98195, U.S.A. olm- [email protected] 3 The Bergius Foundation at the Royal Swedish Academy of Sciences, Box 50017 SE-104 05 Stockholm, Sweden. [email protected]

A phylogenetic study of plastid DNA sequences (ndhF, trnL/F, and rps16) in is presented. In partic- ular, the inclusiveness of Scrophulariaceae sensu APG II is elaborated. Scrophulariaceae in this sense are main- ly a southern hemisphere group, which includes Hemimerideae (including , with a few South American ), Myoporeae, the Central American Leucophylleae (including Capraria), Androya, Aptosimeae, Buddlejeae, Teedieae (including Oftia, Dermatobotrys, and Freylinia), Manuleeae, and chiefly Northern tem- perate Scrophularieae (including Verbascum and Oreosolen). Camptoloma and Phygelius group with Buddlejeae and Teedieae, but without being well resolved to any of these two groups. Antherothamnus is strongly supported as sister taxon to Scrophularieae. African are shown to include Bowkerieae and Charadrophila. There is moderate support for a of putative Asian origin and including Phrymaceae, , , Mazus, Lancea, and chiefly parasitic , to which Brandisia is shown to belong. A novel, strongly supported, clade of taxa earlier assigned to Scrophulariaceae was found. The clade includes Stemodiopsis, Torenia, Micranthemum and probably Picria and has unclear relationships to the rest of Lamiales. This clade possibly represents the tribe Lindernieae, diagnosed by geniculate anterior filaments, usually with a swelling.

KEYWORDS: Lamiales, ndhF, phylogeny, rps16, Scrophulariaceae, trnL/F.

influential classifications for the 19th century concept of INTRODUCTION Scrophulariaceae were those of Bentham (1846), who in Lamiales sensu APG II (The Angiosperm Phylogeny addition recognized Selaginaceae Choisy, Wettstein Group, 2003) constitute a major clade of flowering (1895), who recognized Lentibulariaceae, Plantagin- , with approximately 17,800 species (Judd & al., aceae, and Orobanchaceae as separate families, and 2002) and 23 families, representing 12.3% of eudicot Hallier (1903), who made the broadest circumscription diversity (Stevens, 2001, onwards). They belong to a of the . major asterid clade, the Lamiids (Bremer & al., 2002). Even if doubts sometimes were expressed regarding The monophyly of Lamiales is comparatively uncontro- the naturalness of Scrophulariaceae (e.g., Thieret, 1967; versial and well supported by molecular (e.g., Backlund Barringer, 1984), it was not until the study by Olmstead & al., 1998; Oxelman & al., 1999; Albach & al., 2001; & Reeves (1995) based on plastid ndhF and rbcL gene Bremer & al., 2002) and phytochemical data (Jensen, sequences that the concept of Scrophulariaceae needed a 1992; Scogin, 1992). Morphologically, they typically are revolutionary revision in order to fit within a phyloge- characterized by opposite , sympetalous zygomor- netic framework. Olmstead & Reeves (1995) discovered phic , oligosaccharides, frequent production of 6- two clearly separated consisting of scrophularia- oxygenated flavones, embryos of Onagrad type and cap- ceous taxa. One clade (“scroph I”) included L., sular . Although the majority of the many well- Selago L., Verbascum L. and the type known families in Lamiales are well supported (e.g., Scrophularia L., whereas the other (“scroph II”) includ- Olmstead & al., 2001), the relationships among them are ed L., L., L., L., obscure, despite several recent molecular studies (e.g., Hippuris L., and L. In their study the enig- Olmstead & Reeves, 1995; Backlund & al., 1998; matic woody Paulownia Siebold & Zucc., which was Oxelman & al., 1999; Olmstead & al., 2001). usually classified in Scrophulariaceae but transferred to The concept “Scrophulariae” occurs in Durande, Juss. by Takthajan (1980), and Schlegelia Notions Elém. Bot.: 265. 1782, but the name of the fam- Miq., which was originally placed in Bignoniaceae but ily is conserved as Scrophulariaceae Jussieu. The most argued by Armstrong (1985) to fit better within

411 Oxelman & al. • Disintegration of Scrophulariaceae 54 (2) • May 2005: 411–425

Scrophulariaceae, were both left in uncertain positions accordance with phylogenetic relationships can be pre- within Lamiales. sented. Fischer (2004) has presented a tentative classifi- Several subsequent molecular phylogenetic studies cation of all genera belonging to Scrophulariaceae sensu focusing either on other taxa of Lamiales or more inclu- lato (i.e., approximately in the sense of Hallier, 1903) sive groups (e.g., Soltis & al., 1998; Olmstead & al., into tribes and informal higher groups (“families”). 2000, 2001; Albach & al., 2001; Bremer & al., 2002) The aims of this study are to identify the inclusive- have confirmed the general pattern revealed by Olmstead ness of the “scroph I” clade, sensu Olmstead & Reeves & Reeves (1995). (1995), using plastid DNA sequence data and to infer The parasitic plants that have variously been placed relationships of representatives of major groups of taxa in Scrophulariaceae or Orobanchaceae Vent. have been previously assigned to Scrophulariaceae, for which plas- shown to constitute an additional monophyletic group tid DNA sequences have not been obtained before. (dePamphilis & al., 1997; Wolfe & dePamphilis, 1998; Young & al., 1999). Olmstead & al. (2001) merged the rbcL/ndhF dataset and the data from the plastid rps2 gene that had been useful for the recognition of MATERIAL AND METHODS Orobanchaceae. They also added some other taxa, and Sampling. — Most previous molecular studies were thereby able to identify an additional distinct scro- have used sequences of rbcL and ndhF, but many other phulariaceous clade, . In addition, plastid loci have been introduced as well, including rps2 Halleria L. was shown to group with Stilbaceae Kunth, (dePamphilis & al., 1997), trnL/F intron/spacer region and Mimulus L. did not belong to any of these five clades (Freeman & Scogin, 1999; McDade & Moody, 1999), of ex-Scrophulariaceae taxa. Beardsley & Olmstead rps16 (Wallander & Albert, 2000; Bremer & al., 2002), (2002) identified Mimulus as a member of another clade, matK (Bremer & al., 2002), trnT/F spacer region, and the Phrymaceae. Globularia L. has been demonstrated to trnV spacer (Bremer & al., 2002). We use the ndhF, belong to the Veronicaceae clade sensu Oxelman & al. trnL/F, and rps16 DNA sequence regions, partly because (1999), Olmstead & al. (2001), Kornhall & al. (2001), a substantial number of bulk Lamiales taxa have already and Bremer & al. (2002). been sequenced for these, and partly because previous Olmstead & al. (2001) recognized five distinct phy- studies have indicated that these regions are particularly logenetic lineages composed mainly of taxa previously informative in Lamiales. Initially, we included all assigned to Scrophulariaceae. Nevertheless, several of Lamiales taxa with sequence information from either of their included genera (e.g., Schlegelia, Paulownia, the trnL/F, rps16, or ndhF regions deposited in EMBL/ Mimulus) did not group with any of these five lineages. Genbank as per May 2002. However, because of uneven This fact, together with the restricted sampling (39 of ca. representation, we used an exemplar approach for groups 280 genera in Scrophulariaceae in a traditional sense, whose monophyly is not in question. In addition, we (see Watson & Dallwitz, 1992 onwards), calls for more tried to obtain material for DNA extraction from repre- extensive sampling of genera previously assigned to sentatives of genera that have been suggested to be relat- Scrophulariaceae. ed to Scrophulariaceae sensu APG II. We also included Scrophulariaceae sensu APG II (2003) constitutes representatives from other parts of Scrophulariaceae approximately what Olmstead & Reeves (1995) identi- sensu Hallier and from Stilbaceae. We tried to find addi- fied as the “scroph I” clade, that is, Buddlejaceae K. tional representatives of every taxon, and sequence at Wilh. (see Oxelman & al., 1999 for phylogenetic cir- least one of the three regions to verify the taxonomic cumscription), Manuleeae Benth. (incl. Selagineae, see identity of the sequences. Information on sampled taxa, Kornhall & al., 2001), the two large genera Scrophularia and EMBL/Genbank accession numbers can be found in and Verbascum, plus the tribe Hemimerideae Benth. and the Appendix. Myoporaceae R. Br., including Androya Perrier (Oxel- Sequencing. — Most of the sequencing was per- man & al., 1999; Bremer & al., 2002). The assignment of formed either at the Evolutionary Biology Centre, Hemimerideae to this clade has received only weak sup- Uppsala University, Sweden, or at the Department of port in previous studies. Several taxa that have been sug- Botany, University of Washington, Seattle, U.S.A. Total gested to have affinities with Myoporaceae (e.g., DNA was usually extracted from dried material Capraria L., Anticharis Endl. and Peliostomum E. Mey.) using some variant of a standard CTAB/Chloroform have not been studied by molecular methods. The inclu- extraction protocol, often followed by DNA purification siveness of Manuleeae is not entirely clear, and perhaps using the Qiaquick PCR purification kit (Qiagen). most importantly, a more extensive sampling of taxa that Polymerase chain reactions (PCR) of the targeting have been attributed to Scrophulariaceae is needed regions were performed using combinations of primers before a revised classification of Scrophulariaceae in published in Taberlet & al. (1991), Oxelman & al. (1997,

412 54 (2) • May 2005: 411–425 Oxelman & al. • Disintegration of Scrophulariaceae

1999), Kornhall & al. (2001), and Popp & Oxelman the program Modeltest v. 3.06 (Posada and Crandall, (2001). For trnL/F two new primers were constructed: 1998). We ran the program for 1,000,000 generations, trnLBOC: GGCGRAATYGGTAGACGCTACG, and four parallel chains, and with every 100th saved. trnLBOF-R: CCAGATTTGAACTGGTGACACGAG. In order to detect possible incongruence resulting PCR products were purified using the Qiaquick PCR from analyses of the three regions, a partitioned Bremer purification kit (Qiagen). In some instances, when the support analysis (PBS) was conducted using TreeRot v. 2 products were present as faint bands only, we used nest- (Sorensen, 1999). The are rooted with sequences ed primers in a second round of PCR in order to obtain from Oleaceae Hoffsgg. & Link, which has been shown enough products for sequencing. Automated sequencing to be sister to the rest of Lamiales in previous was performed either on an ABI377 (Applied DNA phylogenies (Savolainen & al., 2000; Bremer & al., Biosystems), or on a MegaBACE 1000 capillary 2001, 2002; Olmstead & al., 2001). machine (Amersham Pharmacia Biotech) using manu- facturer’s protocols. Assembly and alignment. — Sequences were assembled using Sequencher v. 3.1.1 (GeneCodes RESULTS Corporation). The resulting sequences were aligned The number of sequences, previously unreported using the guidelines in Oxelman & al. (1997), and the sequences generated by us, aligned positions, parsimony “simple gap coding” described by Simmons and informative positions and indels, consistency index (CI), Ochotorrena (2000). and retention index (RI), for each region and the com- . — Separate phylogenetic analyses bined analyses are summarized in Table 1. The strict con- were performed for each region initially. Taxa for which sensus tree from 129 most parsimonious trees found in only one region has been sequenced are not presented in the A2 parsimony analysis (at least two sequenced any of the results, unless the relationships found have not regions available) is shown with bootstrap percentages previously been reported in the literature. We made three above branches in Figs. 1–6. Details of the summary tree combined analyses: one where all taxa were included, in Fig. 1 are shown in Figs. 2–6, and referred to in the regardless of completeness (A1), one where at least two discussion of each group. The strict consensus tree from of the three regions were available for each terminal the A3 analysis is shown in Fig. 7 with bootstrap and taxon (A2), and one with taxa where all three regions PBS values. Table 2 shows the bootstrap support for were available (A3). For all analyses, we used maximum comparable nodes in the three analyses. In most cases parsimony as the optimality criterion, and the program there is higher bootstrap support in the A2 and A3 analy- PAUP* ver. 4.0b10 (Swofford, 1999) to find the most ses than in the A1 analysis, and the differences are often parsimonious trees using the TBR search algorithm and considerable. The A2 and A3 analyses had similar sup- 50 random sequence addition replicates, saving a maxi- port levels for comparable groups. mum of ten trees in each. For each analysis, the strict The Bayesian analysis was performed under a gener- consensus tree was calculated from these trees. al time reversible model with a proportion of invariant Bootstrapping was performed with 500 replicates, TBR sites and a gamma distribution, as selected by Modeltest. search, and three random additions per replicate. No The first 100,000 generations were discarded as burn-in. more than 10 trees were saved per random addition repli- The frequencies of different nodes are indicated below cate. branches on the A2 trees in Fig. 1–6. Generally, the par- A Bayesian phylogenetic analysis was performed on simony and the Bayesian analyses are highly congruent, dataset A2 using MrBayes v. 3.0b4 (Huelsenbeck & usually with considerably higher frequencies in the Ronquist, 2001) on computers running Linux. The Bayesian analysis. Cases where the parsimony and appropriateness of different models was evaluated using Bayesian analyses differ include the position of

Table 1. Tree statistics in the form of number of sequences, previously unreported sequences generated by ourselves, aligned positions, parsimony informative sites and indels, consistency index (CI), and retention index (RI), for each region and the combined analyses. Number of Number of Number of Number of Region sequences First time reported aligned positions informative substitutions informative indels CI RI ndhF 235 56 2231 1136 20 0.28 0.62 trnT-F 170 83 1190 501 18 0.46 0.68 rps16 109 106 1076 425 25 0.49 0.60 A3 83 4497 1456 43 0.47 0.52 A2 144 4497 1742 51 0.42 0.62 A1 299 4497 2063 61 0.34 0.63

413 Oxelman & al. • Disintegration of Scrophulariaceae 54 (2) • May 2005: 411–425

89 Scrophulariaceae (Fig. 3) 1. 88 99 Stilbaceae (Fig. 4) 78 1. 1. 98 1. Lindernieae (Fig. 5) 1. 1. 52 ABLLMOPPV (Fig. 6) 1. -1. 79 (Fig. 2) 100 1. 78 1. s.l. .62 100 1. Calceolariaceae 100 Tetrachondraceae 1. Oleaceae

Fig. 1. Simplification of the strict consensus tree from the 129 most parsimonious trees found in the A2 analysis with bootstrap values, when >50%, given above nodes. Bayesian posteriors under a GTR + gamma model are given below branches. Calceolariaceae is represented by a single sequence in this analysis. References to detailed subtrees are given inside parentheses. ABLLMOPPV = , Bignoniaceae, , Lentibulariaceae, Martyniaceae, Orobanchaceae, Pedaliaceae, Phrymaceae, and .

Calceolaria, which in the parsimony analysis is sister Streptocarpus Lindley), and in both cases the rps16 data group to the core of Lamiales (excluding are in conflict with trnL/F and ndhF. We have rechecked Tetrachondraceae and Oleaceae, Fig. 1). The bootstrap the identities of these sequences as well as the alignment, support for this is less than 50%. In the Bayesian analy- and we can find no obvious explanation to this pattern. sis, ends up as to Gesneriaceae There is support both from ndhF (bootstrap) and trnL/F with a posterior probability of 1. Colpias and Alonsoa (bootstrap) for a sister-group relationship between form a monophyletic group with .56 posterior probabili- Sanango and Gesneriaceae. The support for Peltanthera ty, whereas they form a poorly (51% bootstrap frequen- as sister taxon to Sanango and Gesneriaceae is weak cy) supported grade in the parsimony analysis (Fig. 3). The positions of Rehmannia and Paulownia are both Table 2. Comparison of bootstrap support for nodes in poorly resolved within the ABLLMOPPV clade by the the different analyses. Numbers in parentheses are num- ber of taxa in the node. A1 tree not shown. parsimony bootstrap analysis, but form consecutive sis- Group A1 A2 A3 ter groups to Orobanchaceae with 100 % posterior prob- Alonsoa...Verbascum <50 (99) 95 (93) 99 (28) abilities in the Bayesian analysis. Diclis...Hemimeris 98 (5)o 100 (5)o 100 (2)o Alonsoa..Hemimeris <50 (9)o 55 (6)o 53 (6)o Diascia...Nemesia 100 (3)o 100 (3)o 100 (2)o Alonsoa... <50 (13) 72 (10) 72 (6)o DISCUSSION Androya...Verbascum <50 (87) 72 (48) 71 (22) Androya...Peliostomum 76 (17) 75 (9)o 67 (8)o The general structure of the tree in Figure 1 is in Aptosimum...Peliostomum 99 (5)o 100 (3)o 100 (2)o accordance with previously published chloroplast DNA Androya...Leucophyllum 62 (11) 100 (6)o 100 (6)o phylogenies of Lamiales (e.g., Oxelman & al., 1999; frutescens Olmstead & al., 2001; Bremer & al., 2002). With Olea- ...Leucophyllum 75 (10) 100 (5)o 100 (5)o ceae designated as the , Tetrachondraceae frutescens Leucophyllum...Capraria 64 (5)o 100 (2)o 100 (2)o appears as sister to the rest. In agreement with Olmstead Myoporum...Eremophila 94 (5)o 100 (3)o 100 (3)o & al. (2001) and Bremer & al. (2002), there is emerging Bontia...Myoporum 56 (4)o 75 (2)o 73 (2)o support that the bulk of “core” Lamiales does not include Camptoloma...Verbascum 54 (70) 93 (39) 100 (14) Calceolariaceae, Gesneriaceae Rich. & Juss., Sanango Dermatobotrys...Teedia 56 (5)o 90 (3)o 92 (3)o Bunting & Duke, and Peltanthera Benth. The relation- Oftia...Teedia 78 (2)o 88 (2)o 92 (2)o Buddleja...Emorya 81 (7)o 100 (6)o 100 (3)o ships among these are, however, contradictory. The PBS Verbascum...Selago 54 (46) 92 (35) 98 (6)o analysis (Fig. 7) gave conflicting numbers for two nodes Manuleopsis...Selago 92 (36) 84 (26) 93 (3)o (Calceolaria L. as sister group to the rest of Lamiales Antherothamnus...Verbascum 55 (8)o 100 (5)o 100 (3)o including Gesneriaceae, and monophyly of Sanango/ Verbascum...Scrophularia 50 (7)o 98 (4)o 92 (2)o

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100 pubescens 99 1. Basistemon 86 1. 80 Melosperma 1. .99 Monttea 74 Gratiola 1. 98 100 1. Stemodia Gratioleae 98 1. Scoparia 1. Mecardonia 100 Antirrhinum 79 91 1. * 100 1. 1. Gambelia * 1. Maurandella * 100 100 1. 100 Globularia cordif. (1) 100 1. 1. Poskea Globularieae 1. 97 100 Digitalideae 1. 1. Isoplexis obliqua (1) 70 84 100 .97 Penstemon * Cheloneae 1. * .98 100 Russelia Russelieae 1. Tetranema 50 changes

Fig. 2. Phylogram of one of the most parsimonious trees from the A2 analyses with taxa from Plantaginaceae. Bootstrap values, when >50%, are indicated above branches. Bayesian posteriors under a GTR + gamma model are given below branches. * denotes sequences combined from different vouchers. Species epithets and occasionally numbers within parentheses are given to differentiate between DNA samples within genera, when there are more than one (see Appendix). from our data, only trnL/F contributes with positive sup- 2001), and matK and nrDNA ITS sequences (Wolfe & port in the three-gene dataset (Fig. 7). However, both of al., 2002). In our A2 tree, as well as in Wolfe & al., these relationships are supported by rbcL (Oxelman & (2002), and some of the trees obtained by Olmstead & al. al., 1999). Bremer & al. (2002) report strong support for (2001), Russelieae (Russelia Jacq. and Tetranema these relationships, and Jensen (2000) report biochemi- Benth.) forms the sister group to Cheloneae, although in cal evidence that may indicate a relationship between none of the trees the relationship receives strong support. Peltanthera and Gesneriaceae. Our study and Olmstead & al. (2001) also concur In the following discussion, we will present and dis- that Cheloneae and Russelieae form a monophyletic cuss more detailed results from particular groups of the group with Antirrhineae Dumort., Digitalis, Globularia, “core” Lamiales. We use the family classifications of Poskea Vatke, Campylanthus Roth., Hippuris, Olmstead & al. (2001) and APG II and the tribal rank to Callitriche, Plantago, Wallich., and classify the genera studied taxonomically. We also dis- Veronica. This clade consists of several well-delimited cuss some still unsequenced scrophulariaceous taxa and groups, of which several often have been recognized at their putative relationships. the family level (Globulariaceae DC., Hippuridaceae Plantaginaceae (Veronicaceae sensu Olmstead & al. Vest., Callitrichaceae Bercht. & J. Presl. and Plantagina- 2001). This group (Fig. 2) was originally recognized by ceae sensu stricto). Ghebrehiwet & al. (2000) examined Olmstead & Reeves (1995) as “Scroph II” and was cor- Antirrhineae cpDNA relationships in detail, and the sis- roborated with more taxa in Olmstead & al. (2001). The ter-group relationship to the rest of the above-mentioned group is supported in the cpDNA tree (parsimony boot- genera is well established. strap: A2 - 79%; A3 - 91%), and there is also support for Globularia and Poskea form a strongly supported a sister-group relationship to the rest of the core group, which is corroborated by morphological data Lamiales. Within Plantaginaceae, there are several well- (Barringer, 1993). Campylanthus is resolved as sister to supported subgroups. Cheloneae Benth., comprised sole- these, a result which needs corroboration from other ly of New World taxa, have been identified by several data. The proximity of Campylanthus to Digitalideae previous molecular studies, including restriction sites Dumort. has been suggested (see Hjertsson, 2003) and (Wolfe & al., 1997, 2002), the trnL intron (Freeman & has also been substantiated in a phytochemical study by Scogin, 1999), ndhF, rbcL, and rps2 (Olmstead & al., Rønsted & Jensen (2002) and a brief molecular study by

415 Oxelman & al. • Disintegration of Scrophulariaceae 54 (2) • May 2005: 411–425

Alonsoa unilabiata 51 Diclis bambuseti 100 -.56 Diclis petiolaris 100 1. .59 Diclis reptans 1. 100 Hemimeris montana Hemimerideae 70 1. Hemimeris sabulosa Colpias 1. 100 Nemesia cheiranthus 1. 100 1. Nemesia strumosa Androya 100 Bontia 100 Myoporeae 1. Myoporum mauritianum 100 1. Eremophila 75 1. 100 Capraria 1. 1. Leucophyllum frutescens (2) Leucophylleae 100 Anticharis glandulosa 100 1. Peliostomum Aptosimeae 89 1. Aptosimum sp 1. Antherothamnus 100 100 Oreosolen Scrophularia californica 1. 98 1. 99 Scrophularieae 1. Scrophularia peregrina 1. Verbascum arcturus 99 Barthlottia 1. Manuleopsis 100 Chenopodiopsis Pseudoselago ascendens Cromidon 100 92 Microdon dubius 1. 1. Microdon lucidus 98 66 86 1. Selago myrtifolia 81 1. Selago thunb. .97 1. Melanospermum foliosum 100 92 100 1. 1. Melanospermum transvaalense 1. 84 1. Tetraselago .99 96 1. Glumicalyx 97 100 Phyllopodium 1. 1. Polycarena Reyemia 100 80 1. 100 Zaluzianskya glareosa .78 1. Zaluzianskya minima 100 100 Dischisma Limoselleae 60 1. Hebenstretia cordata 1. 1. Trieenea 100 Glekia 1. 85 Manulea cheiranthus 100 73 .99 Manulea glandulosa 93 90 1. Manulea crassifolia 1. 1. 1. Sutera foetida 99 Limosella aquatica 89 1. Limosella grandiflora .98 Lyperia antirrhinoides Jamesbrittenia megadenia 82 Buddleja asiatica Buddleja davidii .93 10056 Buddleja saligna 1. 74 Nicodemia Buddlejeae 52 .91 Emorya suaveolens (2) - .92 59 Gomphostigma 1. 88 Oftia 90 Teedia Teedieae 97 .99 Dermatobotrys * 89 1. .97 Freylinia * .96 Phygelius Camptoloma canariense 50 changes 100 Camptoloma lyperiiflorum 1. Camptoloma rotundifolium Fig. 3. Phylogram of one of the most parsimonious trees from the A2 analysis with taxa from Scrophulariaceae sensu APG II. Bootstrap values, when >50%, are given above nodes (or on the left when this was not possible). Bayesian posteriors under a GTR + gamma model are given below branches. * indicates sequences combined from different vou- chers. Species epithets and occasionally numbers within parentheses are given to differentiate between DNA samples within genera, when there are more than one (see Appendix).

Hjertsson (1997). ported monophyletic groups irrespective of whether Digitalis appears paraphyletic in relation to Isoplexis divergent Callitrichaceae and Plantaginaceae s.s. are Loudon according to ndhF data (results not shown). included. Hemiphragma appears to belong to the According to ITS data (Albach, 2001), Erinus L. is also Plantagineae/Veroniceae clade rather than to Digitalideae part of Digitalideae. (Olmstead & al., 2001). Digitalideae and Globularieae form strongly sup- The least understood group in the former

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ous molecular studies. In addition, tribe Teedieae Benth. 92 Anastrabe 94 Bowkeria verticillata is identified as a member of this clade. In the following 1. .59 Ixianthes retzioides (2) text, each of the tribes is discussed in more detail. Bowkerieae Campylostachys Scrophularieae. — The close relationship 97 100 1. 92 1. Stilbe between Scrophularia and Verbascum has been suggest- 100 Retzia capensis (1) ed by previous molecular studies, as well as similarities 1. .92 99 68 Euthystachys in and embryo characteristics (Thieret, 1967), and 1. .75 Stilbaceae s. Nuxia anatomy (Lersten & Curtis, 1997). The close relation Charadrophila between Oreosolen Hook. f. and Scrophularia (Fig. 3) is Halleria lucida (1) here reported for the first time, but is not surprising from 50 changes a morphological point of view. Both the Himalayan Oreosolen and the closely related Nathaliella B. Fedtsch. Fig. 4. Phylogram of one of the most parsimonious trees from Central agrees well with Scrophularia in flo- from the A2 analysis with representatives from ral morphology and general leaf architecture. The rela- Stilbaceae. Bootstrap values, when >50%, are indicated above the nodes. Bayesian posteriors under a GTR + tionship also makes sense biogeographically, since gamma model are given below branches. Species Scrophularia, Oreosolen, and Verbascum all have main- epithets and occasionally numbers within parentheses ly Northern Hemisphere distributions, as opposed to are given to differentiate between DNA samples within most other Scrophulariaceae. The strong support for a genera, when there are more than one (see Appendix). sister-group relationship between the South African Antherothamnus N. E. Br. and Scrophularia/Oreosolen/ Verbascum is not evident morphologically.The position Scrophulariaceae is probably Gratioleae. The circum- of Antherothamnus is robust to method of analysis, i.e., scription of Gratioleae by Bentham has been discussed neighbor-joining, maximum likelihood, and Bayesian by Wettstein (1895) and Thieret (1967) among others. inference with various evolutionary models (results not Olmstead & al. (2001) reported monophyly of shown), and is supported by all three chloroplast genes Amphianthus Torrey, Aublet, and Gratiola L., (Fig. 7). Similar to Scrophularia, Antherothamnus has a and a weakly supported relationship with Angelonia fully developed staminode, a character that is not found Bonpl. We present data that assign Scoparia L., Stemodia in Manuleeae. Antherothamnus now appears to be a link L., and Mecardonia Ruís & Pavón to Gratioleae. Many between southern African ancestors and Eurasian genera remain to be tested for their relationships with Scrophularieae. [ N.B. The Scrophularia macrantha trnL this group, with many members occurring in Neotropical sequence on EMBL/Genbank is probably a misidentifi- areas. This is even more evident for Angelonieae. The cation, because it groups with Collinsieae with strong close relationship between Angelonia and Basistemon support.] Turcz. previously suggested by Barringer (1993) is cor- Limoselleae. — Manuleeae encompass Selagineae roborated, and these in turn form a strongly supported Wettstein (Selaginaceae Choisy) since the latter are nest- group together with the South American genera Monttea ed within Manuleeae (Kornhall & al., 2001). However, and Melosperma, consecutively followed by a less the unexpected finding that Limosella L. also is nested strongly supported relationship with the Neotropical within this group (Fig. 3, Kornhall & Bremer, 2004) sug- Ourisia Comm. Plantaginaceae deserve more study, in gests that the correct name for this tribe is Limoselleae particular with respect to Gratioleae/Angelonieae. Dumort., for priority reasons (Kornhall & Bremer, 2004). Recently, Plantaginaceae have been studied in more Limosella includes small plants that, as the vernacular detail by Albach & al. (2005). English name, mudworts, indicates, grow in wet areas. Scrophulariaceae sensu APG II (Fig. 3). — Their choice of and the small might have The concept of Scrophulariaceae has changed consider- facilitated a global distribution of the taxon by migrating ably with the greater understanding gained via molecular birds (Darwin, 1872). The inclusion of Selagineae in phylogenetics. Olmstead & al. (2001) and APG II (2003) Limoselleae on molecular evidence is also strongly cor- delimit the family to a clade consisting of Scrophularieae roborated by morphological characters. The “Selagi- Dumort., Manuleeae Benth., Buddlejeae Bartl. (Buddle- neae” gynoecium, with one ovule per locule has appar- jaceae sensu Oxelman & al. 1999), Myoporeae Rchb., ently emanated several times within Limoselleae. Leucophylleae Miers, Aptosimeae Benth. & Hook. f., Maintaining a separate Selagineae and Cronquist’s and Hemimerideae Benth. In this study, we confirm the (1981) view that Selagineae belonged in the Globularia- monophyly of this assemblage (89% bootstrap support in ceae results from accepting the uni-ovulate ovary as a our A2 analysis; 99% in our A3 analysis; Fig. 7). We add cardinal character. In spite of several authors pointing to one to several taxa to each of the tribes relative to previ- the close connection between the two tribes based on

417 Oxelman & al. • Disintegration of Scrophulariaceae 54 (2) • May 2005: 411–425

Picris 86 1. 100 Torenia baillonii (2) 100 1. Torenia polygonioides 98 1. Micranthemum glomeratum 1. Stemodiopsis buchananii 50 changes

Fig. 5. Phylogram of one of the most parsimonious trees from the A2 analysis with taxa from Lindernieae. Bootstrap values, when >50%, indicated above nodes. Bayesian posteriors under a GTR + gamma model are given below bran- ches. Species epithets and occasionally numbers within parentheses are given to differentiate between DNA samples within genera, when there are more than one (see Appendix). other characters (Junell, 1961; Argue, 1993; Hilliard, and Camptoloma are small, and in Phygelius gradually 1994), the two tribes were not united before Kornhall & turning leaf-like. We conclude that Teedia, Oftia, al. (2001). Long branches characterize the basal parts of Dermatobotrys, and Freylinia are best accommodated in Limoselleae, and the relations between Jamesbrittenia, Teedieae, whereas at the present it cannot be determined Limosella and Lyperia are not totally clear. Kornhall & which relationship Phygelius and Camptoloma have to Bremer (2004) elaborate in more detail on the interpreta- Teedieae and Buddlejeae. Teedieae, Buddlejeae, tion of morphological features of Limosella in relation to Camptoloma, Phygelius, Manuleeae, and Scrophularieae the chloroplast phylogeny found here. form a very strongly supported monophyletic group Buddlejeae. — The results presented here corrob- based on chloroplast DNA sequences. orate the circumscription of Buddlejeae by Oxelman & Myoporeae. — With small variations, Myopora- al. (1999). Wallick & al. (2000) have, with more exten- ceae have generally been circumscribed as a mostly sive sampling of Buddleja and trnL intron sequences, Southern Hemispheric taxon including Myoporum Sol., further strengthened the suggestion that Buddleja might Eremophila R. Br., and Bontia L. Oftia, here considered be paraphyletic. to belong to Teedieae (see above), has sometimes been Teedieae. — There is strong support for a mono- considered to belong here (e.g., Wettstein, 1895), but phyletic group, Teedieae, consisting of the genera Oftia Dahlgren & Rao (1971) rejected this based on morpho- Adans., Teedia Rudolphi, Dermatobotrys Bolus, and logical evidence. The monophyly of Myoporum, Freylinia Colla. This group in turn forms a strongly sup- Eremophila, and Bontia is here strongly supported, a ported monophyletic group consisting also of the like- result consistent with the palynological data presented by wise woody Buddlejeae, Phygelius Mey., and Niezgoda & Tomb (1975) and anatomical data presented Camptoloma Benth. Wallick & al. (2000) reported a sis- by Carlquist & Hoekman (1986). ter-group relationship between Buddlejeae and Teedieae, Leucophylleae have been suggested to be related to but they included only Teedia and Oftia in their study, so Myoporeae (palynological data: Niezgoda & Tomb, their results are in agreement with ours. Phygelius and, in 1975; anatomical data: Carlquist & Hoekman, 1986; particular, Camptoloma are poorly resolved with respect molecular data: Oxelman & al., 1999; Olmstead & al., to Teedieae and Buddlejeae. Barringer (1993) segregated 2001). The close relationship between Leucophyllum the African woody taxa Teedieae, Freylinieae, and Bonpl. and Capraria L., both with Latin American dis- Bowkerieae from Cheloneae where Bentham originally tribution, reported by Niezgoda & Tomb (1975) based on placed them. Freylinieae sensu Barringer included also palynological evidence, and by Lersten & Curtis (2001) Phygelius, Antherothamnus, and Manuleopsis Thell., a based on anatomical evidence is here corroborated. [N.B. relationship that is not supported by the present study. The Leucophyllum minus trnL sequence deposited on Barringer did not consider the epiphytic Dermatobotrys EMBL/Genbank (AF034878) does not form a mono- in this context. Morphologically, Buddlejeae, Teedieae, phyletic group with our Leucophyllum frutescens and Phygelius all have anthers with separate thecae, sequence. Instead the trnL sequence labeled Pedicularis whereas the anther thecae of Camptoloma are confluent procera (AF034877) groups with L. frutescens. The (as in Manuleeae and Scrophularieae). It is difficult to identities of the L. minus and P. procera sequences need determine the polarity of this character. Possibly, the confirmation.] leafy typical for Oftia, Teedia, Karrfalt & Tomb (1983) and Lersten & Beaman Dermatobotrys, and Freylinia could be interpreted as a (1998) claimed homology between the oil cavities found synapomorphy for this group, as the in Buddlejeae in several Leucophyllum species and those found in the

418 54 (2) • May 2005: 411–425 Oxelman & al. • Disintegration of Scrophulariaceae

100 Barleria 86 1. Chlamydacanthus 91 1. Crossandra * 1. Thunbergia alata Pinguicula Lentibulariaceae Acanthaceae 52 Elytraria * 68 99 Kigelia Bignoniaceae 1. Tecoma stans (2) Martynia 100 Martyniaceae 1. Proboscidea Stachytarpheta dichotoma (2) Verbenaceae .82 Schlegelia parviflora 2 100 Berendtia 100 52 1. Hemichaena Phrymaceae 100 1. Phryma 1. 1. Mimulus 97 Brandisia 58 100 1. Cyclocheilon .92 1. Lindenbergia philippinensis 100 Lancea

59 1. Mazus stachydifolius Orobanchaceae 1. -1. -1. Rehmannia * Paulownia 100 dichotoma Lamiaceae 1. Lamium Sesamum Pedaliaceae 50 changes

Fig. 6. Phylogram of one of the most parsimonious trees from the A2 analyses with representatives for the families Acanthaceae, Bignoniaceae, Lamiaceae, Lentibulariaceae, Martyniaceae, Orobanchaceae, Pedaliaceae, Phrymaceae and Verbenaceae. Bootstrap values, when >50%, are indicated above nodes. Bayesian posteriors under a GTR + gamma model are given below branches. * denotes sequences combined from different vouchers. Species epithets and occa- sionally numbers within parentheses are given to differentiate between DNA samples within genera, when there are more than one (see Appendix). three Myoporeae genera, a hypothesis that is consistent pollen (Erdtman 1952; Niezgoda & Tomb, 1975; Punt, with the results presented here. However, Lersten & 1980). Thus, these features may be synapomorphies for Beaman (1998) found no cavities in L. revolutum Rzed or this group (“Myoporaceae”). in Eremogeton grandiflorus Standl. & L. O. Williams, Hemimerideae. — The chiefly South African which is generally considered to be closely related to tribe Hemimerideae (including Alonsoa) is supported by Leucophyllum. More detailed molecular phylogenetic our data (70% bootstrap) and conforms well to the cir- studies on Leucophyllum, Eremogeton, and the little cumscription reviewed in Steiner (1996), except that the known Faxonanthus Greenm. from Guatemala, are war- chloroplast DNA sequence data support an inclusion of ranted. Colpias E. Mey. Steiner & Whitehead (1996) argued that There is a strongly supported sister-group relation Colpias rather should belong Bowkerieae, based on a between Myoporeae and Leucophylleae and between basic chromosome number of x = 20, a tubular corolla, that group and the Madagascar genus Androya (100% in and a staminode corresponding to a fifth fertile , both the A2 and the A3 analyses). characters that are shared with Bowkerieae. The amphi- Aptosimeae. — The circumscription of Atlantic Alonsoa groups weakly with Diclis and Aptosimeae, characterized by alternate leaves and a Hemimeris. All have a basic chromosome number of x = dilated corolla tube, is one of the very few that seems to 7 (Steiner, 1996). The South American species of have been stable over the history of Scrophulariaceae Alonsoa form a monophyletic group based on ndhF classification. Bentham included Aptosimum, Pelio- sequences (results not shown). Their shared ancestry is stomum (sometimes included in Aptosimum), and further supported by having 2n = 56 instead of 2n = 28 in Anticharis, and has been followed by Wettstein among the South African taxa. Thus, the interpretation of the others. Monophyly of Aptosimeae is strongly supported absence of oil secretion in South American species of by our chloroplast DNA sequences, and a sister group Alonsoa as a synapomorphy (Steiner, 1996) is reinforced. relation with Androya, Leucophylleae, and Myoporeae is Whether the origin of Alonsoa predates the breakup of moderately supported. All have 3-colpate, diporate Gondwanaland needs to be further investigated. Nemesia

419 Oxelman & al. • Disintegration of Scrophulariaceae 54 (2) • May 2005: 411–425

53 Alonsoa unilabiata 2,-1,0 100 Diclis reptans 12,25,10 Hemimeris sabulosa 70 2,0,0 Colpias 0,1,2 100 Diascia 17,2,0 Nemesia strumosa Androya 100 76 Bontia 100 10,14,7 0,0,1 Myoporum mauritianum 99 100 4,2,4 Eremophila 1,3,5 65 12,7,10 100 Capraria 1,2,1 14,4,2 Leucophyllum frutescens * 100 Aptosimum sp. 18,4,6 Peliostomum 70 100 Antherothamnus 9,5,1 92 Scrophularia californica 1,1,1 98 5,0,0 Verbascum arcturus 4,2,1 93 Manuleopsis 73 5,0,1 Selago thunbergii Scrophulariaceeae 100 6,-1,-2 Limosella * 100 Buddleja davidii 4,10,7 10,8,3 95 Emorya suaveolens (2) 2,0,1 Nicodemia 87 Camptoloma rotundifolium 92 1,0,1 Oftia 92 0,1,1 Teedia 95 3,-1,1 Dermatobotrys * 3,0,2 Phygelius 89 Anastrabe 0,0,4 Ixianthes retzioides (2) 98 100 Campylostachys 94 3,0,2 Stilbe 3,0,3 100 1,1,1 Retzia capensis (1) 100 66 15,7,6 Euthystachys 4,5,2 1,0,0 Nuxia

Charadrophila Stilbaceae Halleria lucida (1) 100 Barleria 100 25,12,0 Chlamydacanthus Acanthaceae 15,1,8 Thunbergia alata 100 Berendtia 100 19,7,7 Hemichaena 100 14,4,6 Phryma Phrymaceae 59 11,3,4 Mimulus -2,2,1 100 Brandisia Orobanchaceae 63 26,3,6 Lindenbergia 2,1,0 100 Lancea 88 1,0,0 39,9,20 Mazus stachydifolius 2,0,-1 Rehmannia * 8,0,-1 Paulownia 69 Catalpa Bignoniaceae 5,0,-1 Lamium Lamiaceae Proboscidea Martyniaceae 1,0,0 Schlegelia 100 Stachytarpheta dichotoma (2) 4,6,8 Lantana * 100 99 Micranthemum 29,11,17 Torenia (2) Lindernieae 4,5,4 Stemodiopsis Sesamum Pedaliaceae 100 Angelonia pubescens 54 100 14,4,11 Basistemon 66 4,1,6 80 Melosperma 6,0,-3 2,0,0 -1,1,3 Monttea 75 Ourisia 5,2,-2 93 Gratiola 100 6,4,-1 Stemodia 21,20,9 Scoparia 91 Antirrhinum 4,0,3 99 100 Campylanthus 100 4,3,4 99 13,7,0 100 Globularia cordifolia (1) 52 11,6,9 Poskea 11,7,11 7,4,2 Plantaginaceae 82 1,1,0 Isoplexis Chelone obliqua (1) 4,2,-1 100 Russelia 18,1,10 Tetranema Peltanthera 0,1,0 82 Sanango 6,6,-5 Streptocarpus * Calceolaria Ligustrum 100 Polypremum 44,6,22 Tetrachondra patagonica

Fig. 7. Strict consensus tree from the A3 analysis. Parsimony bootstrap values, when >50%, are indicated above nodes. Below nodes the values from the PBS analysis are indicated in the following order: ndhF, trnL/F and rps16. *denotes sequences combined from different vouchers. Species epithets and occasionally numbers within parentheses are given to differentiate between DNA samples within genera, when there are more than one (see Appendix).

Vent. is here shown to be closely related with Diascia. in the folding of lateral corolla lobes in bud and cited Steiner (1996) showed that these taxa share the basic unpublished chloroplast rps2 gene sequences as evidence chromosome number x = 9, but emphasized differences for Nemesia belonging outside Hemimerideae. Our data

420 54 (2) • May 2005: 411–425 Oxelman & al. • Disintegration of Scrophulariaceae

reject this, and instead favor the similarities in the androecium pointed out by Hilliard & Burtt (1984) as ACKNOWLEDGEMENTS being homologous for Nemesia and Diascia. Reija Dufva, Per Erixon, Nahid Heidari, Jan-Eric Mattsson, Stilbaceae. — The circumscription of this family and Pat Reeves are gratefully acknowledged for invaluable work continues to expand (Fig. 4). Bremer & al. (1994) pre- in the lab. Numerous persons helped us by providing plant materi- sented molecular data that supported the close relation- al, we are especially grateful to the help from Dirk Albach, Fanny ship between Retzia Thunb. and Stilbe Berg. proposed by Astholm, Mandy Balkwill, Mark W. Chase, Søren R. Jensen, Goldblatt & Keating (1976) on morphological grounds. Pieter Maas, Veronica Mayer, Heidi Meudt, Mats Thulin, Andrea Oxelman & al. (1999) showed that Nuxia Comm., previ- Wolfe, the staff at Uppsala Botanical Garden, and the curators of ously classified in Buddlejaceae, also belong here. These the GB, S, and UPS herbaria. Kåre Bremer and Mats Thulin are findings are supported by phytochemical data (Damtoft gratefully acknowledged for comments on earlier versions of the & al., 1984; Frederiksen & al., 1999). Olmstead & al. manuscript. The study was financed by grants from the Swedish (2001) added Halleria, and in this study we show that the Research Council to BO and BB, respectively. chloroplast DNA phylogeny supports putting the tribe Bowkerieae (Bowkeria, Anastrabe, and Ixianthes; Barringer, 1993) and Charadrophila Marloth here. Charadrophila capensis is a rare plant found in shade on LITERATURE CITED permanent wet cliffs in the Cape Province. Its position Albach, D. C. 2001. Paraphyly of Veronica (Veronicaceae; has been disputed ever since Marloth’s original descrip- Scrophulariaceae): evidence from the internal transcribed tion (Marloth, 1898). He placed it in Scrophulariaceae, spacer (ITS) sequences of nuclear ribosomal DNA. J. Pl. Res. 114: 9–18. but Engler, in a note to the original description placed it Albach, D. C., Meudt, H., & Oxelman, B. 2005. Piecing in Gesneriaceae, which it resembles superficially. Weber together the “new” Plantaginaceae. Amer. J. Bot. 92: (1989) followed Marloth and placed it in Scrophulari- 297–315. aceae arguing for a close relationship with Alonsoa. Its Albach, D. C., Soltis, P. S., Soltis, D. S. & Olmstead, R. G. position in Stilbaceae was unexpected, and detailed stud- 2001. Phylogenetic analysis of based on ies are needed in order to establish homology hypotheses sequences of four genes. Ann. Missouri Bot. Gard. 88: in this expanded circumscription of Stilbaceae. 163–212. Argue, C. L. 1993. Pollen morphology in the Selagineae, Lindernieae. — A novel strongly supported clade Manuleae (Scrophulariaceae), and selected Globulari- of taxa earlier assigned to Scrophulariaceae was found aceae, and its taxonomic significance. Amer. J. Bot. 80: that includes Stemodiopsis Engl., Torenia L., Micranthe- 723–733. mum Desf., and probably Picria Lour. (= Conobea) and Armstrong, J. E. 1985. The delimitation of Bignoniaceae and has unclear relationships to the rest of Lamiales (Fig. 4). Scrophulariaceae based on floral anatomy, and the place- This clade probably represents the tribe Lindernieae, ment of problem genera. Amer. J. Bot. 72: 755–766. diagnosed by geniculate anterior filaments, usually with Backlund, M., Oxelman, B. & Bremer, B. 1998. Phylogenetic relationships within the Gentianales based a basal swelling. If this character proves to be synapo- on ndhF and rbcL sequences, with a particular reference to morphous, then for example also Lindernia All., the . Amer. J. Bot. 87: 1029–1043. Craterostigma Hochst., Crepidorhopolon E. Fischer, Barringer, K. A. 1984. Seed morphology and the classification Hartliella E. Fischer, and Artanema D. Don. may belong of the Scrophulariaceae. Amer. J. Bot. 71 (5, pt. 2): 156. here. Neither Stemodiopsis, Picria, or Micranthemum [Abstr.] possess the characteristic basal swelling of the base of Barringer, K. A. 1993. Five new tribes in the the filament, but like Stemodiopsis, they have geniculate Scrophulariaceae. Novon 3: 15–17. Beardsley, P. M. & Olmstead, R. G. 2002. Redefining or curved anterior filaments/staminodes. Lindernieae Phrymaceae: the placement of Mimulus, tribe Mimuleae, have mostly been classified in Gratioleae (Fischer, and Phryma. Amer. J. Bot. 89: 1093–1102. 1997), but have a pretty distinct morphology (e.g., alve- Bentham, G. 1846. Ordo CXLII Scrophulariaceae. In: de olate endosperm of a certain type; Fischer, 1992). Candolle, A. de (ed.), Prodromus Systematis Naturalis Orobanchaceae, Phrymaceae and relatives. Regni Vegetabilis, vol. 10. Victoris Masson, Paris. — There is moderate support for a clade including Bremer, K., Backlund, A., Sennblad, B., Swenson, U., Phrymaceae, Paulownia, Rehmannia Libosch., Mazus Andreasen, K., Hjertsson, M., Lundberg, J., Backlund, M. & Bremer, B. 2001. A phylogenetic analysis of 100+ Lour., Lancea Hook. f. & Thomson, and chiefly parasitic genera and 50+ families of Euasterids based on morpho- Orobanchaceae, to which Brandisia Hook. f. & logical and molecular data with notes on possible higher Thomson, previously not known to be parasitic, is shown level morphological synapomorphies. Pl. Syst. Evol. 229: to belong (Fig. 6). The inclusion of Lancea and Mazus in 137–169. Phrymaceae, as advocated by Beardsley & Olmstead Bremer, B., Bremer, K., Heidari, N., Erixon, P., Olmstead, (2002), is not supported here. R. G., Anderberg, A. A., Källersjö, M. &

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Barkhordarian, E. 2002. Phylogenetics of Asterids based Campylanthus (Scrophulariaceae). Edinburgh J. Bot. 60: on 3 coding and 3 non–coding chloroplast DNA markers 131–174. and the utility of non–coding DNA at higher taxonomic Huelsenbeck, J. P. & Ronquist, F. 2001. MRBAYES, levels. Molec. Phylogen. Evol. 24: 274–301. Bayesian inference of phylogeny. Bioinformatics 17: Bremer, B., Olmstead, R. G., Struwe, L. & Sweere, J. A. 754–755. 1994. rbcL sequences support exclusion of Retzia, Jensen, S. R. 1992. Systematic implications of the distribution Desfontainia, and Nicodemia from the Gentianales. Pl. of iridoids and other chemical–compounds in the Syst. Evol. 190: 213–230. Loganiaceae and other families of the Asteridae. Ann. Carlquist, S. & Hoekman, D. A. 1986. anatomy of the Missouri Bot. Gard. 79: 284–302. Myoporaceae: ecological and systematic considerations. Jensen, S. R. 2000. Chemical relationships of Polypremum Aliso 11: 317–334. procumbens, Tetrachondra hamiltonii and Peltanthera Cronquist, A. 1981. An Integrated System of Classification of floribunda. Biochem. Syst. Ecol. 28: 45–51. Flowering Plants. Columbia Univ. Press, New York. Judd, W. S., Campbell, C. S., Kellogg, E. A., Stevens, P. F. & Dahlgren, R. & Rao, V. S. 1971. Genus Oftia Adans. and its Donoghue, M. J. 2002. Plant Systematics: A Phylogenetic systematic position. Bot. Not. 124: 451–472. Approach, ed. 2. Sinauer Associates, Sunderland, Damtoft, S., Franzyk, H., Jensen, S. R. & Nielsen, B. J. Massachusetts. 1984. Iridioids and verbascoids in Retzia. Junell, S. 1961. Ovarian morphology and taxonomical position 34: 239–243. of Selagineae. Svensk Bot. Tidskr. 55: 168–192. Darwin, C. 1872. The Origin of Species by Means of Natural Karrfalt, E. E. & Tomb, A. S. 1983. Air spaces, secretory cav- Selection. John Murray, London. ities, and the relationship between Leucophylleae dePamphilis, C. W., Young, N. D. & Wolfe, A. D. 1997. (Scrophulariaceae) and Myoporaceae. Syst. Bot. 8: 29–32. Evolution of plastid gene rps2 in a lineage of hemipara- Kornhall, P., Heidari, N. & Bremer, B. 2001. Selagineae and sitic and holoparasitic plants: many losses of photosynthe- Manuleeae, two tribes or one? Phylogenetic studies in the sis and complex patterns of rate variation. Proc. Natl. Scrophulariaceae. Pl. Syst. Evol. 228: 199–218. Acad. Sci. U.S.A. 94: 7367–7372. Kornhall, P. & Bremer, B. 2004. New circumscription of the Erdtman, G. (ed.). 1952. Angiosperms. Pollen Morphology tribe Limoselleae (Scrophulariaceae) that includes the taxa and Plant . Almqvist & Wiksell, Stockholm. of the tribe Manuleeae. Bot. J. Linn. Soc. 146: 453–467. Fischer, E. 1992. Systematik der afrikanischen Lindernieae Lersten, N. R. & Beaman, J. M. 1998. First report of oil cav- (Scrophulariaceae). Trop. Subtrop. Pflanzenwelt 81: ities in Scrophulariaceae and reinvestigation of air spaces 1–365. in leaves of Leucophyllum frutescens. Amer. J. Bot. 85: Fischer, E. 1997. Revision of the genus Stemodiopsis Engl. 1646–1649. (Scrophulariaceae-Gratioloideae). Bot. Jahrb. Syst. 119: Lersten, N. R. & Curtis, J. D. 1997. Anatomy and distribution 305–326. of foliar idioblasts in Scrophularia and Verbascum Fischer, E. 2004. Scrophulariaceae. Pp 333–432 in: Kadereit, (Scrophulariaceae). Amer. J. Bot. 84: 1638–1645. J. W. (ed.), The Families and Genera of Flowering Plants, Lersten, N. R. & Curtis, J. D. 2001. Idioblasts and other vol. VII. Flowering Plants. Dicotyledons: Lamiales unusual internal foliar secretory structures in (except Acanthaceae including Avicenniaceae). Springer, Scrophulariaceae. Pl. Syst. Evol. 227: 63–73. Berlin. Marloth, R. 1898. Charadrophila Marloth nov. gen. Bot. Frederiksen, L. B., Damtoft, S. & Jensen, S. R. 1999. Jahrb. Syst. 26: 358–359. Biosynthesis of iridoids lacking C-10 and the chemotaxo- McDade, L. A. & Moody, M. L. 1999. Phylogenetic relation- nomic implications of their distribution. Phytochemistry ships among Acanthaceae: evidence from noncoding trnL- 52: 1409–1420. trnF chloroplast DNA sequences. Amer. J. Bot. 86: 70–80. Freeman, C. E. & Scogin, R. 1999. Potential utility of chloro- Niezgoda, C. J. & Tomb, A. S. 1975. Systematic palynology plast trnL (UAA) gene intron sequences for inferring phy- of tribe Leucophylleae (Scrophulariaceae) and selected logeny in Scrophulariaceae. Aliso 18: 141–159. Myoporaceae. Pollen & Spores 17: 495–516. Ghebrehiwet, M., Bremer, B. & Thulin, M. 2000. Phylogeny Olmstead, R. G., dePamphilis, C. W., Wolfe, A. D., Young, of the tribe Antirrhineae (Scrophulariaceae) based on mor- N. D., Elisens, W. J. & Reeves, P. A. 2001. Disintegration phological and ndhF sequence data. Pl. Syst. Evol. 220: of the Scrophulariaceae. Amer. J. Bot. 88: 348–361. 223–239. Olmstead, R. G., Jansen, R. K., Kim, K.-J. & Wagstaff, S. J. Goldblatt, P. & Keating, R. C. 1976. Chromosome cytology, 2000. The phylogeny of the Asteridae s.l. based on chloro- pollen structure, and relationship of Retzia capensis. Ann. plast ndhF sequences. Molec. Phylogen. Evol. 16: 96–112. Missouri Bot. Gard. 63: 321–325. Olmstead, R. G. & Reeves, P. A. 1995. Evidence for the poly- Hallier, H. 1903. Ueber die Abgrenzung und Verwandtschaft phyly of the Scrophulariaceae based on chloroplast rbcL der einzelnen Sippen. Bull. Herb. Boissier 2: 181–207. and ndhF sequences. Ann. Missouri Bot. Gard. 82: Hilliard, O. 1994. The Manuleae: a Tribe of Scrophulariaceae. 176–193. Edinburgh Univ. Press, Edinburgh. Oxelman, B., Backlund, M. & Bremer, B. 1999. Hilliard, O. & Burtt, B. L. 1984. A revision of Diascia section Relationships of the Buddlejaceae s.l. investigated using Racemosae. J. S. African Bot. 50: 269–340. parsimony jackknife and branch support analysis of Hjertsson, M. 1997. Systematics of Lindenbergia and chloroplast ndhF and rbcL sequence data. Syst. Bot. 24: Campylanthus (Scrophulariaceae). Comprehens. Summ. 164–182. Uppsala Diss. Fac. Sci. Technol. 331: 46. Oxelman, B., Lidén, M. & Berglund, D. 1997. Chloroplast Hjertsson, M. 2003. Revision of the disjunct genus rps16 intron phylogeny of the tribe Silenae

422 54 (2) • May 2005: 411–425 Oxelman & al. • Disintegration of Scrophulariaceae

(Caryophyllaceae). Pl. Syst. Evol. 206: 393–410. data. Amer. J. Bot. 87: 1827–1841. Popp, M. & Oxelman, B. 2001. Inferring the history of the Wallick, K. P., Elisens, W. J., Molvray, M. & Kores, P. J. polyploid Silene aegaea (Caryophyllaceae) using plastid 2000. A phylogenetic study of Buddlejaceae s.s. inferred and homoeologous nuclear DNA sequences. Molec. from the trnL-F region of cpDNA. Amer. J. Bot. 87 (6, Phylogen. Evol. 20: 474–481. suppl.): 183. [Abstr.] Posada, D. & Crandall, K. A. 1998. MODELTEST, testing Watson, L. & Dallwitz, M. J. 1992. The Families of the model of DNA substitution. Bioinformatics 14: Flowering Plants: Descriptions, Illustrations, 817–818. Identification, and Information Retrieval. http://biodiver- Punt, W. 1980. Pollen morphology. Pp. 162–191 in: Leeuwen- sity.uno.edu/ delta/angio/ berg, A. J. M. (ed.), Engler and Prantl’s Die natürlichen Weber, A. 1989. Family position and conjectural affinities of Pflanzenfamilien, Angiospermae: Ordnung Gentianales, Charadrophila capensis Marloth. Bot. Jahrb. Syst. 111: Fam. Loganiaceae, vol. 28b(1). Duncker and Humblot, 87–119. Berlin. Wettstein, R. von. 1895. Scrophulariaceae. Pp. 39–107 in: Rønsted, N. & Jensen, S. R. 2002. Iridoid glucosides and caf- Engler, A. & Prantl, K. (eds.), Die natürlichen feoyl phenylethanoid from Campylanthus sal- Pflanzenfamilien, vol. 4(3b). Wilhelm Engelmann, saloides and Campylanthus glaber. Biochem. Syst. Ecol. Leipzig. 30: 1091–1095. Wolfe, A. D., Elisens, W., Watson, L. & dePamphilis, C. W. Savolainen, V., Fay, M. F., Albach, D. C., Backlund, A., 1997. Using restriction–site variation of PCR-amplified Bank, M. van der, Cameron, K. M., Johnson, S. A., cpDNA genes for phylogenetic analysis of tribe Cheloneae Lledó, M. D., Pintaud, J.-C., Powell, M., Sheahan, M. (Scrophulariaceae). Amer. J. Bot. 84: 555–564. C., Soltis, D. E., Weston, P., Whitten, W. M., Wurdack, Wolfe, A. D., Datwyler, S. L. & Randle, C. 2002. A phyloge- K. J. & Chase, M. W. 2000. Phylogeny of the : a netic and biogeographic analysis of the Cheloneae nearly complete familial analysis based on rbcL gene (Scrophulariaceae) based on ITS and matK sequence data. sequences. Kew Bull. 55: 257–309. Syst. Bot. 27: 138–148. Scogin, R. 1992. The distribution of acetoside among Wolfe, A. D. & dePamphilis, C. W. 1998. The effect of angiosperms. Biochem. Syst. Ecol. 20: 477–480. relaxed functional constraints on the photosynthetic gene Simmons, M. P. & Ochoterena, H. 2000. Gaps as characters rbcL in photosynthetic and nonphotosynthetic parasitic in sequence-based phylogenetic analyses. Syst. Biol. 49: plants. Molec. Biol. Evol. 15: 1243–1258. 369–381. Young, N. D., Steiner, K. E. & dePamphilis, C. W. 1999. The Soltis, D. E., Soltis, P. S., Mort, M. E., Chase, M. W., evolution of in Scrophulariaceae/ Orobanchac- Savolainen, V., Hoot, S. B. & Morton, C. M. 1998. eae: plastid gene sequences refute an evolutionary transi- Inferring complex phylogenies using parsimony: an tion series. Ann. Missouri Bot. Gard. 86: 876–893. empirical approach using three large DNA datasets for angiosperms. Syst. Biol. 47: 32–42. Sorensen, M. D. 1999. TreeRot. Boston Univ., Boston. Steiner, K. E. 1996. Chromosome numbers and relationships in tribe Hemimerideae (Scrophulariaceae). Syst. Bot. 21: 63–76. Steiner, K. E. & Whitehead, V. B. 1996. The consequences of specialization for in a rare South African , Ixianthes retzioides (Scrophulariaceae). Pl. Syst. Evol. 201: 131–138. Stevens, P. F. 2001. Angiosperm Phylogeny Website. http://www.mobot.org/MOBOT/Research/APweb/ welcome.html Swofford, D. L. 1999. PAUP, Phylogenetic Analysis Using Parsimony (*and Other Methods). Sinauer Associates, Sunderland, Massachusetts. Taberlet, P., Gielly, L., Pautou, G. & Bouvet, J. 1991. Universal primers for amplification of three non–coding regions of chloroplast DNA. Pl. Molec. Biol. 17: 1105–1109. Takhtajan, A. L. 1980. Outline of the classification of - ing plants (Magnoliophyta). Bot. Rev. 46: 225–359. The Angiosperm Phylogeny Group. 2003. An update of the angiosperm phylogeny group classification for the orders and families of flowering plants: APG II. Bot. J. Linn. Soc. 141: 399–436. Thieret, J. W. 1967. Supraspecific classification in the Scrophulariaceae: a review. Sida 3: 87–106. Wallander, E. & Albert, V. A. 2000. Phylogeny and classifi- cation of Oleaceae based on rps16 and trnL-F sequence

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Appendix. Taxon table with EMBL/GenBank accession numbers. An asterisk after the accession number indicates sequences here reported for the first time. Voucher collector and number, with acronym in parentheses, are given for those DNA samples that have previously not been reported in the literature. New sequences with no voucher information are based on the same DNA sample as those previously deposited on GenBank. Taxon; Voucher; Accession number ndhF; Accession number trnL; Accession number rps16. reptans L.; L36391; -; -. Alonsoa auriculata Diels; Molau 1629 (GB); AJ617587*; -; -. Kuntze; Astholm 9 (GB); AJ617612*; -; -. Alonsoa peduncularis (Kuntze) Wettst.; Wall 15/9 -41 (S); AJ619549*, AJ619550*; -. Alonsoa unilabiata Ruiz & Pav.ex Steud.; AF188184; AJ608620*; AJ609217*. Aloysia gratissima (Gill & Hook) Tronc.; AF130154; -; -. Amphianthus pusillus Torr.; AF123674; -; -. Amphitecna apiculata A.H. Gentry; AF102624; -; -. corsicum Jord. & Fourr.; AJ245815; -; -. Anastrabe integerrima E. Mey. ex Benth.; Wells 3588 (S); AJ619551*, AJ619552*; AJ608633*; AJ609216*. Kornhall 88 (UPS); -; -; AJ621104*. Androya decaryi H. Perrier; AF027276; AJ608619*; AJ609215*. Angelonia angustifolia Benth.; Lundin 1019 (S); AJ617588*; -; -. Angelonia pubescens Benth.; AF123675; AJ608618*; AJ609214*. Anisacanthus thurberi A. Grey; -; AF034864; -. Antherothamnus pearsonii N.E. Br.; AJ401392; AJ608617*; AJ609213*. Anticharis glandulosa Aschers.; Thulin et al. 8268 (UPS); AJ619553*; -; AJ609212*. Anticharis linearis (Benth.) Hochst. ex Asch.; Wanntorp & Wanntorp 963 (S); -; -; AJ609211*. L.; L36392; AJ608634*; AJ609218*. Aptosimum indivisum Burch.; Bremer 3508 (UPS); AJ617594*; -; -. Aptosimum sp.; AF123676; AJ608607*; AJ609210*. Arrabidaea pubescens (L.) A.H.Gentry; AF102625; -; -. procumbens Mill.; AJ250380; -; -. alba Blume; -; AY008820; -. Avicennia ger- minans L.; -; AY008819; -. Avicennia marina ssp. australasica (Walp.) J.Everett; -; AY008821; -. Bacopa caroliniana Robinson; AF123677; -; -. Barleria prionitis L.; U12653; AF063118; -. Barthlottia madagascariensis Fischer; AJ401438; AJ401443, AJ401444; -. Bartsia alpina L.; AF123678; -; -. Basistemon klugii Barringer; Plowman & al. 11387 (GB); AJ619554*, AJ619555*; AJ608616*; AJ609209*. Berendtia laevigata B.L. Rob. & Greenm.; Pringle 6294 (S); -; AJ608615*; AJ609208*. Bontia daphnoides L.; Cult. Pacific Trop. Bot. Gard. #711537004; AJ617613*; AJ608614*; AJ609219*. Bowkeria cymosa MacOwan; Balkwill 1510 (S); -; -; AJ609207*. Bowkeria verticillata Druce; Bremer 3767 (UPS); AJ617589*; -; AJ609206*. Brandisia hancei Hook. f.; Kingdon-Ward 318 (UPS); AJ619577*, AJ619578*; AJ608613*; AJ609205*. Buddleja asiatica Lour.; AF027277; AF380858; -. Buddleja davidii Franch.; AF130143; AJ608612*; AJ609204*. Buddleja indica Lam.; L36405; AJ608581*; AJ609158*. Buddleja saligna Willd.; AJ401396; AJ401397; AJ609203*. liniflora Salisb.; AF482605; AF482533; -. Calceolaria mexicana Benth.; AF123679; AJ608611*; AJ609202*. Callicarpa dichotoma Raeusch.; L36395; AF363665; -. Callicarpa japonica Thunb.; AF130148; -; -. Callitriche hermaphroditica (1) L.; L47329; -; -. Callitriche her- maphroditica (2) L.; L36396; -; -. L.; L47330; -; -. Campsis radicans Seem.; AF130144; -; -. Camptoloma canariense (Webb & Berth) Hilliard; AJ401398, AJ401399; AJ401445, AJ401449. Camptoloma lyperiiflorum (Vatke) Hilliard; AJ401401; AJ296514; -. Camptoloma rotundifolium Benth.; AJ401431, AJ401432; AJ401450, AJ296515; AJ609201*. Campylanthus salsoloides Roth.; Bremer 3864 (UPS); AJ619556*; AJ608600*; AJ609200*. Campylostachys cernua Kunth.; Kornhall 135 (UPS); AJ617611*; AJ608621*; AJ609199*. Capraria biflora L.; Cul. Utrecht Bot. Gard. 92GRO151g; AJ617610*; AJ608608*; AJ609198*. Caryopteris bicolor (Roxb.ex Hardw.) D.J.Mabberley; U78680; -; -. Caryopteris divaricata Maxim.; U78679; -; -. integra A. Gray; -; AF118794; -. Catalpa speciosa Warder ex Engelm.; L36397; AJ608599*; AJ609197*. minus L.; AJ250381; -; -. Charadrophila capensis Marloth; Kornhall 131 (UPS); AJ617590*; AJ608622*; AJ609196*. Chelone obliqua (1) L.; AF123680; AF034875; AJ609220*. Chelone obliqua (2) L.; AJ245814; -; -. Chenopodiopsis retrorsa Hilliard; AJ401421; AJ296502; -. Chilopsis linearis Sweet; -; AF034889; -. Chlamydacanthus lindavianus H.Winkler; AJ249408; AJ608598*; AJ609195*. trichotomum C.P.Thunberg ex A.Murray; AF130146; -; -. Codonanthe carnosa (Gardner) Hanst.; -; AY047147; -. Codonanthe digna Wiehler; AF130156; -; -. Collinsia grandiflora Dougl.ex Lindl.; AF188182; -; -. Collinsia heterophylla R.Grah.; -; AF034873; -. Colpias mollis E. Mey.; Jensen IOK-19-99 (C); AJ617591*; AJ608623*; AJ609194*. Conobea multifida (Michx.) Benth.; Fisher 1929 (UPS); -; AJ608597*; -. Cordylanthus kingii S.Wats.; -; AF118795; -. Crescentia portoricensis Britton; AF102627; -; -. Cromidon decumbens (Thunb.) Hilliard; AJ401403; AJ296503; -. Crossandra nilotica Oliver; U12656; -; -. Crossandra pungens Lindau; -; AF061825; -. Picria fel-terrae Lour.; Liang 63174 (GB; -; AJ608609*; AJ609193*. Cybistax donnell-smithii () Seibert; AF102628; -; -. Cyclocheilon somaliense Oliver; Thulin 8364 (UPS); -; AJ608624*; AJ609192*. Cydista aequinoctialis Miers; AF102629; -; -. muralis Gaertn.Mey. & Scherb.; AJ250382; -; -. hawaiensis C.B.Clarke; AF130159; -; -. Dermatobotrys saundersii Bolus; Kornhall 136 (UPS); AJ617592*; -; AJ609191*. Dermatobotrys saundersii Bolus; Jensen IOK-19-99 (C); -; AJ608596*; -. Dermatobotrys saundersii Bolus; -; AJ608635*; AJ609221*. Diascia capsularis Benth.; AJ617593; AJ608595*; AJ609190*. Diclis bambuseti R.E. Fries; Jonsell 2212 (UPS); AJ617614*; AJ608639*; . Diclis petio- laris Benth.; Wanntorp & Wanntorp 955 (S); -; AJ608594*; AJ609189*. Diclis reptans Benth.; Kornhall 104 (UPS; AJ619557*; AJ608593*; AJ609188*. Lam.; L36399; -; -. Digitalis purpurea L.; AF130150; AF034871; -. Dischisma ciliatum Choisy; AJ401412; AJ296501; -. Drymonia serrulata (Jacq.) Mart.; -; AY047152; -. Drymonia strigosa Wiehler; AF130158; -; -. scaber Ruíz & Pav.; AF102630; -; -. Elytraria cre- mata Vahl.; U12657; -; -. Elytraria imbricata Vahl; -; AF061819; -. Emorya suaveolens (1) Torr.; -; AF327415; -. Emorya suaveolens (2) Torr.; AF027278; AJ608625*; AJ609187*. Eremophila bignoniflora F. Muell.; Olmstead 794 (WTU); AJ617609*; AJ608592*; AJ609186*. Euthystachys abbreviata A. DC.; AF147715; AJ608626*; AJ609185*. Freylinia lanceolata (L. f.) G. Don; Rourke 1991 (S); -; -; AJ609184*. Freylinia tropica S. Moore; AJ401402; -; -. fruticosa J.F.Gmel.; AJ250383; -; -. Nutt. syn w Galvezia sp.; AJ250384; -; -. Gambelia speciosa A.Gray; -; AF118796; -. Glekia krebsiana (Benth.) Hilliard; AJ401422; AJ296519; -. Globularia cordifolia (1) L.; AF027282; AJ430930; AJ431055. Globularia cordifolia (2) L.; AF124557; -; -. Globularia nudicaulis L.; AF123681; -; -. Glumicalyx flanaganii (Hiern) Hilliard; AJ401413; AJ296497; -. Gmelina hystrix Schult.ex Kurz; U78692; -; -. Gomphostigma scoparioides Turcz.; AF027279; -; AJ609183*. Gratiola pilosa Michx.; AF188183; AJ608591*; AJ609182*. Halleria lucida (1) L.; AF188185; AJ608590*; AJ609181*. Halleria lucida (2) L.; Williams 2316 (S); -; -; AJ609180*. Hebenstretia cordata L.; AJ401414; AJ296499; -. Hebenstretia dentata L.; AF027285; -; -. Hemichaena fruticosa Benth.; Castillo 1745 (S); AJ617595*; AJ608632*; AJ609179*. Hemimeris montana L.f.; Wall 18/9-1938 (GB); -; -; AJ609230*. Hemimeris montana L.f.; Goldblatt 2787 (S); AJ617596*; -; AJ609178*. Hemimeris sabulosa L.f.; AF123682; AJ608638*; AJ609227*. Hemiphragma heterophyllum Wall.; AF123683; -; -. Hippuris vulgaris L.; L36401; -; -. Holmskioldia sanguinea Retz.; U78693; -; -. Howelliella ovata (Eastw.) Rothm.; AJ250385; -; -. Isoplexis canariensis Steud.; Thulin 9945 (UPS); AJ617597*; AJ608589*; AJ609177*. Ixianthes retzioides (1) Benth.; Adamson 4298 (BOL); -; -; AJ609176*. Ixianthes retzioides (2) Benth.; Wolfe 97.118; AJ617598*; AJ608636*; AJ609222*. Jacaranda sparrei A.H.Gentry; AF102631; -; -. Jamesbrittenia dissecta Kuntze; AJ401435,AJ401436; -; -. Jamesbrittenia filicaulis (Benth.) Hilliard; AJ401439; -; -. Jamesbrittenia megadenia Hilliard; AJ401404; AJ296511; -. Jasminum fluminense Vell.; -; AF102445; -. Jasminum mesnyi Hance; AF130162; -; -. Jovellana sp.; AF123684; -; -. Justicia carnea Lindl.; AF130155; -; -. Justicia longii Hilsenb.; -; AF063135; -. Keckiella antirrhinoides (Benth.) Straw; -; AF034876; -. elatine (L.) Dumort.; AJ245816; -; -. Kickxia gracilis D.A.Sutton; AJ245817; -; -. Kigelia africana Benth.; AF102632; AF034880; -. Lamium purpureum L.; U78694; AJ608588*; AJ609175*. Lancea tibetica Hook.f. & Thoms.; Ho et al. 158 (E); AJ617599*; AJ608610*; AJ609174*. Lantana camara L.; -; -; AF225294. Lantana camara L.; -; AY008824; -. Lantana horrida H.B. & K.; AF130152; -; -. Leucocarpus alatus D.Don; -; -; AJ609173*. Leucocarpus perfoliatus Benth.; AJ617600*; -; -. Leucophyllum frutescens (1) I.M.Johnst.; -; AF380873; -. Leucophyllum frutescens (2) I.M.Johnst.; AF123685; -; AJ609172*. Leucophyllum frutescens (3) I.M.Johnst.; Schallert 18738 (S); -; -; AJ609171*. Leucophyllum minus A.Gray; -; AF034878; -. Ligustrum vulgare L.; AF130164; AF231848; AF225257. Limosella aquatica L.; Dvorak 1556 (UPS); AJ619558*; AJ608587*; -. Limosella grandiflora Benth.; AJ550552; -; AJ609170*. amethystea (Lam.) Hoffm. & Link; AJ250386; -; -. Linaria canadensis Dum.-Cours.; -; AF034867; -. Lindenbergia indica (L.) Vatke; AF027286; -; -. Lindenbergia philippinensis Benth.; AF123686; AJ608586*; AJ609169*. Lyperia antirrhinoides (L.f.) Hilliard; AJ401405; AJ296521; -. Lyperia tristis Benth.; AJ401406; -; -. acerifolia (Pennell) Elisens; -; AF118797; -. Macfadyena unguis-cati (L.) A.H Gentry; AF102633; -; -. Manulea cheiranthus L.; AJ401418 AJ401419; AJ401446 AJ401452. Manulea

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Appendix (continued). Taxon; Voucher; Accession number ndhF; Accession number trnL; Accession number rps16. crassifolia Benth.; AJ401428 AJ401429; AJ401453 AJ296518. Manulea glandulosa Philips; AJ401395; AJ296520; -. Manulea tomentosa Hiern; AJ401394; -; -. Manuleopsis dinterii Thell.; AJ401410; AJ401454, AJ296500; AJ609168*. Martynia annua L.; AF190906; AF067065; -. Maurandella antirrhiniflora (Willd.) Rothm.; AJ250387; -; -. Maurandella antirrhiniflora (Willd.) Rothm.; -; AF118798; -. scandens A.Gray; AJ245818; -; -. Mazus reptans N.E. Br.; -; AF479004; -. Mazus stachydifolius Maxim.; Ulanova & Basargin 6936 (S); AJ619559*; AJ607432*, AJ607433*; AJ609167*. Mecardonia flagellaris (C. & S.) Rossow; Iversen et al. 87345 (UPS); AJ617601*; AJ608627*; AJ609166*. Melanospermum foliosum (Benth.) Hilliard; AJ401415; AJ401456, AJ296507; -. Melanospermum transvaalense (Hiern) Hilliard; AJ401426, AJ401427; AJ401455, AJ296508; -. Melasma scabrum Berg.; AF123687; -; -. Melosperma andicola (Gill.) Benth.; Wall 21/12-1946 (GB); AJ617602*; AJ608585*; AJ609165*. Micranthemum glomeratum (Chapm.) Shinners; Tehler et al. 22 (S); AJ617603*; AJ608584*; AJ609164*. Micranthemum umbrosum (J.F.Gmel.) Blake; -; AF034879; -. Microdon dubius (L.) Hilliard; AJ401411; AJ296491; -. Microdon lucidus Choisy; AJ401416; AJ296492; -. Mimulus ringens L.; AF188186; AJ608606*; AJ609163*. orontium (L.) Raf.; AJ250388; -; -. Mohavea breviflora Coville; -; -; AJ609223*. (DC.) A. A. Heller; AJ250389; -; -. Monttea chilensis Gay; Frödin 246 (UPS); AJ617604*; AJ608583*; AJ609162*. Myoporum insulare R.Br.; -; -; AJ299258. Myoporum mauritianum A.DC.; L36403; AJ608582*; AJ609161*. Myoporum montanum R.Br.; -; AJ296513; -. Nematanthus longipes DC.; AF130157; -; -. Nematanthus strigillosus; -; AY047148; -. Nemesia cheiranthus E. Mey. ex Benth.; Kornhall 19 (UPS); AJ617605*; -; AJ609160*. Nemesia strumosa Benth.; AF123688; AJ608631*; AJ609159*. Nothochelone nemorosa (Douglas ex Lindl.) Straw; -; AF034874; -. Nuxia sp.; AF027280; AJ608605*; AJ609157*. Nyctanthes arbor-tristis L.; AF130161; -; -. Oftia africana Bocq.ex Baill.; Bremer 1152 (S); AJ617606*; AJ60858*; AJ609156*. Olea europaea L.; AF027288; AF231866; -. Olea europaea L.; AF130163; -; -. Ophiocolea floribunda (Boj.ex Lindl.) H.Perrier; AF102634; -; -. Oreosolen unguiculatus Hemsl.; Paulsen 1921 (S); AJ617607*; -; AJ609155*. Oroxylum indicum (L.) Kurz; AF102635; -; -. Ourisia poeppigii Benth.; Meudt & Lopes 13 (TEX); AJ619560*, AJ619561*, AJ619562*; AJ608579*; AJ609154*. Paliavana prasinata Benth. & Hook.f.; AF130160; -; -. Pandorea jasminoides Schum; AF102636; -; -. (Thunb.) Steud.; L36406; AJ608578*; AJ609153*. Pedicularis foliosa L.; AF123689; -; -. Pedicularis procera A.Gray; -; AF034877; -. Peliostomum leucorrhizum E. Mey. ex Benth.; Skarpe 1976-12-16 (UPS); AJ619563*, AJ619564*; AJ608577*; AJ609152*. Peltanthera floribunda Benth; AF027281; AJ608576*; AJ609151*. Penstemon centranthifolius Benth.; -; AF034883; -. Penstemon sp.; Oxelman 2338 (WTU); -; AJ619565*; --. Petitia domingensis Jacq.; U78697; -; -. Phryma leptostachya L.; AJ617586*; AF478988/ AJ430928; AJ609150*. Phygelius capensis E.Mey.ex Benth.; Oxelman 2339 (UPS); AJ617608*; AJ608575*; AJ609149*. Phyla incisa Small; AF130153; -; -. Phyllopodium cuneifolium (L.f.) Benth; AJ401430; AJ401457,AJ296496; -. Pinguicula sp.; Erixon 54 (UPS); -; AJ608604*; AJ609148*. Plantago lanceolata L.; AF130151; -; -. Podranea ricasoliana Sprague; AF102637; -; -. Polycarena formosa Hilliard; AJ401423; AJ296515; -. Polypremum procumbens L.; AJ011986; AJ431063; AJ430938. Poskea socotrana (I.B.Balf.) G.Taylor; Thulin & Gifri 8670 (UPS); AJ617585*; AJ608574*; AJ609147*. Prasium majus L.; U78700; -; -. Premna microphylla Turcz.; U78701; -; -. Proboscidea louisianica (Mill.) Thell.; AF123690; AJ608573*; AJ609146*. Pseuderanthemum alatum Radlk.; -; AF063130; -. Pseudoselago ascendens (E.Mey.) Hilliard; AJ401433; AJ296495; -. Pseudoselago serrata (P.J.Bergius) Hilliard; AJ401440; -; -. Pseudoselago subglabra Hilliard; -; AJ296498; -. Radermachera frondosa Chun & How; AF102638; -; -. Ramonda myconii (L.) Rchb.; -; -; -. Rehmannia angulata Hemsl.; Hedberg 29.9.1943 (UPS); AJ619566*; -; AJ609145*. N.E.Brown; -; AJ608572*; -. Retzia capensis (1) Thunb.; AF027289; AJ608628*; AJ609144*. Retzia capensis (2) Thunb.; AF147716; -; -. Reyemia chasmantiiflora Hilliard; AJ401425; AJ296505; -. atrosanguineum (Zucc.) Rothm.; AJ250390; -; -. Ruellia californica I.M.Johnst.; -; AF063115; -. Ruellia ciliata Hornem.; U12664; -; -. Russelia retrorsa Greene; Penell Aug 1995 (S); AJ619567*; AJ608571*; AJ609143*. Salvia divinorum Epling & Jativa; U78703; -; -. Sanango sp.; AF027283; AJ608603*; AJ609142*. Sarmienta scandens Pers.; -; -; -. Schlegelia parviflora (1) (Oerst.) Monach.; -; AY008825; -. Schlegelia parviflora (2) (Oerst.) Monach.; L36410; AJ608570*; AJ609141*. pterosperma A.Br.; AJ250391; -; -. Scoparia dulcis L.; Ryding 683 (UPS); AJ619568*, AJ619569*; AJ608569*; AJ609140*. Scrophularia californica Cham. & Schlecht.; L36411; AF118802; AJ609224*. Scrophularia macrantha Greene ex Stiefelhagen; -; AF034865; -. Scrophularia nodosa L.; -; AF118803; -. Scrophularia peregrina L.; Julin 23.4.1985 (UPS); -; AJ608568*; AJ609139*. Selago canescens L.f.; L36412; AJ608637*; AJ609225*. Selago corymbosa L.; -; AJ401458, AJ296494; -. Selago myrtifolia Hilliard; AJ401420*; AJ296500; -. Selago setulosa Rolfe; -; AJ296493; -. Sesamum indicum L.; L36413; AJ608602*; AJ609226*. Seymeria pectinata Pursh; AF123691; -; -. Sinningia schiffneri Fritsch; -; -; -. Stachytarpheta dichotoma (1) Vahl; -; AY008823; -. Stachytarpheta dichotoma (2) Vahl; L36414; AJ608567*; AJ299259. Stemodia glabra Oerst.; Nordenstam & Anderberg 967 (S); AJ617584*; AJ608566*; AJ609138*. Stemodiopsis buchananii Skan; Iversen & Martinsson 89169 (UPS); AJ619570*, AJ619571*, AJ619572*; AJ608565*; AJ609137*. Stemodiopsis humilis Skan; Ngoni 177 (P); -; -; AJ609229*. Stilbe albiflora E.Mey.; AF027287; AJ608629*; AJ609136*. Streptocarpus caulescens Vatke; Erixon 35 (UPS); -; AJ608601*; AJ609135*. Streptocarpus holstii; L36415; -; -. Strobilopsis wrightii Hilliard & B.L.Burtt; AJ401441; -; -. Sutera foetida Roth; AJ401407, AJ401408; AJ296510; -. Sutera hispida (Thunb.) Druce; AJ550566, AJ550567; -; -. Sutera patriotica Hiern; AJ401393; -; -. Syringa emodi Wall.; AF130228; -; -. Syringa vulgaris L.; -; AF231882; -. Tecoma stans (1) Juss.; -; AY008826; -. Tecoma stans (2) Juss.; AF130145; AF034888; -. Tecomaria capensis (Thunb.) Spach; -; AY008827; -. Tectona grandis L.f; AF027284; - ; -. Teedia lucida (Ait.) Rud.; Bremer 3545 (UPS); AJ617582*; AJ608561*; AJ609127*. Tetrachondra hamiltonii Petrie; -; -; AJ609134*. Tetrachondra patagonica Skottsb.; AF027272; AJ430939; AJ609133*. Tetraclea coulteri A.Gray; AF130147; -; -. Tetranema mexicanum Benth.; AF123692; AJ608630*; AJ609132*. Tetraselago longituba (Rolfe) Hilliard & Burtt; AJ401417; AJ296506; -. Teucrium fruticans L.; U78686; -; -. Thunbergia alata Bojer ex Sims; U12667; AJ608564*; AJ609131*. Thunbergia erecta T.Anders.; -; -; AJ609228*. Torenia baillonii (1) Godefroy ex Andre; Oxelman 2367 (UPS); -; AJ608563*; -. Torenia baillonii (2) Godefroy ex Andre; Olmstead 98-52 (WTU); AJ617583*; AJ608562*; AJ609130*. Torenia polygonioides Benth.; Klackenberg & Lundin 624 (S); AJ619573*, AJ619574*; -; AJ609129*. Trieenea glutinosa (Schltr.) Hilliard; AJ401400; AJ401459,. AJ296516. Uncarina grandidieri (Baill.) Stapf; -; AF363667; -. Verbascum arcturus L.; AJ619575*, AJ619576*; AJ401460, AJ296522; AJ609128*. Verbascum nigrum L.; -; AF118804; -. Verbascum thapsus L.; L36417; -; -. Verbena bonariensis L.; L36418; -; -. Verbena urticifolia L.; -; AY008822; -. Veronica catenata Pennell; L36419; -; -. Vitex agnus-castus L.; AF130149; -; -. Zaluzianskya benthamiana Walp.; AJ401409; -; -. Zaluzianskya glareosa Hilliard & Burtt; AJ401424; AJ401448, AJ296504; -. Zaluzianskya katharinae Hiern; AF123693; -; -. Zaluzianskya minima (Hiern) Hilliard; AJ401437; AJ401447; - Combined sequences: Codonanthe digna/carnosa/-; AF130156/AY047147/-. Collinsia grandiflora/heterophylla/-; AF188182/AF034873/-. Crossandra nilotica/pungens/-; U12656/AF061825/-. Dermatobotrys saundersii; AJ617592*; AJ608635*; AJ609191*. Drymonia strigosa/serrulata/strigosa; AY047152/AY047152/ AF130158. Elytraria cremata/imbricata/-; U12657/AF061819/-. Freylinia tropica/-/lanceolata; AJ401402/-/AJ609184*. Gambelia speciosa/speciosa/-; AJ250384/AF118796/-. Lantana horrida; AF130152; AY008824; AF225294. Leucophyllum frutescens; AF123685; AF380873; AJ609172*. Limosella grandiflora/aquatica/grandiflora; AJ550552/AJ608587*/AJ609170*. Maurandella antirrhiniflora/antirrhiniflora/-; AJ250387/AF118798/-. Mohavea con- fertiflora/-/breviflora; AJ250389/-/AJ609223*. Nematanthus longipes/strigillosus/-; AF130157/AY047148/-. Penstemon sp./centrantifolius/-; AJ619565*/AF034883/-. Rehmannia angulata/elata/angulata; AJ619566*/AJ608572*/AJ609145*. Streptocarpus holstii/caulescens/caulescens; L36415/ AJ608601*/AJ609135*.

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